Chromatin immunoprecipitation (ChIP) assays were performed using the MAGnify Chromatin Immunoprecipitation System (Thermo Fisher Scientific, 49–2024) as previously described (26 (link)). IgG antibodies were included in the kit and anti-HSF1 (Abcam ChIP Grade, AB2923), anti-MYC-tag (Cell Signaling, 2278), anti-histone H3-acetyl K27 (Abcam ChIP Grade, AB4729), anti-histone H3-mono methyl K4 (Abcam ChIP Grade, AB8895) antibodies were used according to the manufacturer’s instructions. The set of primers used for ChIP allows amplification of target regions including HSF1 binding sites at intron 2 and at the 5′UTR region of the locus. FOXO3 exon 3 region was used as a control (sequences are listed in ‘Additional Material’).
Protocol detail
ChIP Assay of HSF1, MYC, and Histone Marks
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Antibodies
Assays
Binding sites
Chromatin immunoprecipitation
Exon
Histone h3
Igg antibodies
Intron
Primers
Corresponding Organization : Gastroenterology Hospital "Saverio de Bellis"
Other organizations : Sapienza University of Rome
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Variable analysis
independent variables
- Antibody used for chromatin immunoprecipitation (ChIP): IgG antibodies, anti-HSF1, anti-MYC-tag, anti-histone H3-acetyl K27, anti-histone H3-mono methyl K4
- Enrichment of target regions including HSF1 binding sites at intron 2 and at the 5'UTR region of the locus, as measured by ChIP-qPCR
- FOXO3 exon 3 region used as a control
- Positive control: IgG antibodies included in the kit
- Negative control: Not explicitly mentioned
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